Aggregation of a-synuclein (aS) plays an important but still poorly understood role in the pathogenesis of Parkinson's disease (PD). The normal function of aS remains unknown, but the protein binds to phospholipid membranes and is believed to regulate synaptic vesicle pool size, vesicle recycling, neurotransmitter transport and release, and synaptic plasticity. Since the discovery of the link between aS and PD, there has been great interest in identifying aS interaction partners that influence either the pathogenic or normal roles of the protein. In the past few years, a growing number of proteins, polymers, and small molecules have been reported to bind to aS and alter its aggregation kinetics and/or its lipid-associated functions. Among these are two other members of the synuclein family, B-synuclein (BS) and y-synuclein (yS). Covalent modifications also affect aS aggregation and function. However, the mechanisms by which these various binding interactions and modifications influence aS behavior are not well understood. We have shown that residual structure in free aS may play an important role in mediating the intermolecular interactions that precede amyloid fibril formation by this protein. We hypothesize that upon covalent modification or partner interactions, aS undergoes conformational changes that underlie the consequent effects on the aggregation or normal functions of the protein. We therefore propose to characterize, at high resolution, the structural changes that occur in aS as a function of its modifications and its interactions with different partners, with an initial focus on (BS, yS, histones, HSP70, PLD2, copper and other metals and polycations. We also propose to elucidate in detail the structure of (BS and yS in their free and lipid-bound states to clarify why aS exhibits different self-assembly behavior from these close relatives. Finally, we plan to test our conclusions regarding the role of structural changes in aS by introducing rationally designed mutants, collaboratively, into yeast and fly models of aS toxicity and function. The proposed studies are focused on improving our understanding of the molecular mechanisms underlying PD and may suggest strategies for developing new PD therapeutics. The results may have broader implications for understanding and treating other amyloid diseases, including Alzheimer's disease and the prion diseases.